Safety Profiling of AAV5-Id3 gene therapy using rabbits

Abstract

Purpose : Previously, we reported expression of Inhibitor of differentiation (Id) genes in the cornea and its role in corneal fibrosis in vitro. Our ongoing in vivo studies found remarkably inhibited of corneal haze/fibrosis in vivo in rabbits by Id3 gene therapy given via AAV5. This study sought to evaluate safety and toxicity (acute/short- and long-term) of AAV5-Id3 gene therapy in vivo using rabbits.

Methods : New Zealand White rabbits were used for defining toxicity and safety of AAV5-Id3. Rabbits were divided into 3 groups: Group-1 eyes received 75μL balanced salt solution (BSS) topically (n=12), Group-2 eyes received AAV5-naked vector titer (75μL; 8.23x10¹²μg/mL; n=12), and Group-3 eyes received AAV5-Id3 titer (75μL; 5.63x10¹²μg/mL; n=12). Slit-lamp biomicroscopy, stereo biomicroscopy, specular biomicroscopy, pachymetry, Schirmer test, Intra ocular pressure (IOP), and Modified MacDonald-Shadduck tests were used to evaluate response of AAV5-Id3 in eyes of live rabbits. Corneal tissues were used to study toxicity and safety at cellular and molecular levels using TUNEL assay, Inflammatory biomarkers (CD11b), and other commercial kits. Student’s t-test, analysis of variance (ANOVA) and Bonferroni adjustment for repeated measures were used for statistical analysis.

Results : In situ slit-lamp clinical exams in the eyes of live rabbits did not find clinically-relevant adverse symptoms such as edema, chemosis, ocular discharge, corneal haze, corneal neovascularization, tear flow or IOP in all 3 groups. Also, Fantes, Draize, and Modified MacDonald-Shadduck tests detected no significant abnormalities in the eyes of 3 groups. Corneal tissues collected 2-weeks after AAV5-Id3 gene transfer displayed no noticeable changes in corneal morphology in H&E or abnormal corneal cell death in TUNEL assay. The molecular analyses of 2-week time and ocular safety/toxicity studies after 3-month times are underway.

Conclusions : AAV5-Id3 gene therapy appears safe for rabbit eyes in vivo.

This is a 2020 ARVO Annual Meeting abstract.